Disulfiram modulated ROS-MAPK and NFκB pathways and targeted breast cancer cells with cancer stem cell-like properties

Background:

Previous studies indicate that disulfiram (DS), an anti-alcoholism drug, is cytotoxic to cancer cell lines and reverses anticancer drug resistance. Cancer stem cells (CSCs) are the major cause of chemoresistance leading to the failure of cancer chemotherapy. This study intended to examine the effect of DS on breast cancer stem cells (BCSCs).

Methods:

The effect of DS on BC cell lines and BCSCs was determined by MTT, western blot, CSCs culture and CSCs marker analysis.

Results:

Disulfiram was highly toxic to BC cell lines in vitro in a copper (Cu)-dependent manner. In Cu-containing medium (1 μ), the IC₅₀ concentrations of DS in BC cell lines were 200–500 n. Disulfiram/copper significantly enhanced (3.7–15.5-fold) cytotoxicity of paclitaxel (PAC). Combination index isobologram analysis demonstrated a synergistic effect between DS/Cu and PAC. The increased Bax and Bcl2 protein expression ratio indicated that intrinsic apoptotic pathway may be involved in DS/Cu-induced apoptosis. Clonogenic assay showed DS/Cu-inhibited clonogenicity of BC cells. Mammosphere formation and the ALDH1^+VE and CD24^Low/CD44^High CSCs population in mammospheres were significantly inhibited by exposure to DS/Cu for 24 h. Disulfiram/copper induced reactive oxygen species (ROS) generation and activated its downstream apoptosis-related cJun N-terminal kinase and p38 MAPK pathways. Meanwhile, the constitutive NFκB activity in BC cell lines was inhibited by DS/Cu.

Conclusion:

Disulfiram/copper inhibited BCSCs and enhanced cytotoxicity of PAC in BC cell lines. This may be caused by simultaneous induction of ROS and inhibition of NFκB.     

戒酒硫的比色测定

根据戒酒硫和铜离子形成有色络合物的性质,比色法测定了其含量。探讨并选择了最佳实验条件。本法具有灵敏度高、简便、快速、准确度高、重现性好等特点。     

Comparison of disulfiram and placebo in treatment of alcohol dependence of adolescents

About 50% of alcoholic patients relapse within 3 months of treatment. Previous studies have suggested that disulfiram may help to prevent such relapse. The aim of our study was to assess the efficacy and safety of long-term disulfiram treatment in alcohol dependence of adolescents. In this double-blind, placebo-controlled study we recruited 26 adolescents, aged 16 - 19 years, with chronic or episodic alcohol dependence. Patients were allocated treatment randomly with disulfiram (200 mg daily) or placebo for 90 days. Patients were assessed on the day treatment started and on days 30 and 90 by interview, self-report, questionnaire and laboratory screening. Patients were classified as abstinent, relapsing or non-attending. Time to first treatment failure (relapse or non-attendance) was the primary outcome measure. The disulfiram (n =13) and placebo (n = 13) groups were well matched in terms of baseline demographic and alcohol-related variables. Thirteen disulfiram-treated and 13 placebo-treated patients completed the treatment phase; seven (1 vs. 6) relapsed, five (3 vs. 2) refused to continue treatment, three (1 vs. 2) had concurrent illness and two (1 vs. 1) had adverse side effects. At the end of treatment, seven disulfiram-treated and two placebotreated patients had been abstinent continuously (p= 0.0063). Mean cumulative abstinence duration was significantly greater in the disulfiram group than in the placebo group [68.5 (SD 37.5) vs. 29.7 (19.0) days; p =0.012]. Apart from occasional diarrhoea, there was no difference in side effects between groups. In some cases, disulfiram may be an effective and well-tolerated pharmacological adjunct to psychosocial and behavioural treatment programmes for treatment of adolescent alcohol-dependent patients. [Niederhofer H, Staffen W. Comparison of disulfiram and placebo in treatment of alcohol dependence of adolescents. Drug Alcohol Rev 2003;22:295 - 297]     

双硫仑联合顺铂对三阴性乳腺癌MDA-MB-231细胞的增殖抑制作用及其机制

目的：观察双硫仑（DSF）和顺铂（CDDP）联合使用对三阴性乳腺癌（TNBC） MDA-MB-231细胞的增殖抑制作用,并探讨其可能机制。方法：将MDA-MB-231细胞分为空白对照组、2μmol·L^-1DSF组、60μmol·L^-1CDDP组和联合组（60μmol·L^-1CDDP+1、2和4μmol·L^-1DSF）。MTT法检测各组MDA-MB-231细胞的存活率,流式细胞术检测各组MDA-MB-231细胞凋亡率,流式细胞术检测各组MDA-MB-231细胞中活性氧（ROS）水平;电感耦合等离子体质谱仪（ICP-MS）检测各组MDA-MB-231细胞中铂（Pt）水平。结果：MTT法和流式细胞术检测,作用72 h后,与60 μmol·L^-1 CDDP组比较,联合1组（60 μmol·L^-1CDDP+1 μmol·L^-1DSF）、联合2组（60 μmol·L^-1CDDP+2 μmol·L^-1DSF）和联合3组（60 μmol·L^-1CDDP+4 μmol·L^-1 DSF） MDA-MB-231细胞存活率降低（P<0.05）。作用24 h后,与60 μmol·L^-1 CDDP组和2 μmol·L^-1 DSF组比较,联合组（60 μmol·L^-1CDDP+2 μmol·L^-1 DSF） MDA-MB-231细胞凋亡率升高（P<0.05）。流式细胞术检测,与CDDP组比较,联合组（60 μmol·L^-1CDDP+2 μmol·L^-1 DSF） MDA-MB-231细胞中ROS水平升高（P<0.05）。ICP-MS检测,与CDDP组比较,联合组（20 μmol·L^-1CDDP+0.625 μmol·L^-1 DSF） TNBC MDA-MB-231细胞中Pt水平升高（P<0.05）。结论：DSF与CDDP联合使用能明显抑制MDA-MB-231细胞的增殖,其可能机制是通过提高MDA-MB-231细胞中ROS水平和Pt水平抑制肿瘤细胞生长。     

Allergic "contact" dermatitis from disulfiram implants.

Two cases of allergic "contact" dermatitis to implanted disulfiram (tetraethythiuramdisulfide or TETD) pellets are described. This side effect has never been reported before. Other local complications from disulfiram implantations are reviewed.     

双硫仑联合铜通过调控视神经萎缩蛋白1诱导舌癌细胞凋亡

目的 探讨视神经萎缩蛋白1（OPA1）在双硫仑联合铜（DSF-Cu）诱导舌癌细胞凋亡中的作用。 方法 用不同浓度的DSF-Cu干预舌癌细胞Cal-27,MTT法检测细胞增殖活性,Annexin V-FITC/PI染色结合流式细胞仪检测细胞凋亡,荧光素酶发光法检测ATP含量,Western blot法检测OPA1蛋白表达水平。 结果 DSF-Cu能抑制舌癌细胞的增殖活性,促进细胞凋亡的发生,减少细胞ATP含量,并呈浓度依赖性。Western blot结果示低浓度DSF-Cu对OPA1蛋白表达无明显影响,而中、高浓度的DSF-Cu显著抑制OPA1蛋白表达水平。 结论 DSF-Cu可诱导舌癌细胞凋亡,其机制可能是通过抑制OPA1表达。     

Cu/Zn superoxide dismutase plays a role in angiogenesis.

Endothelial cells produce oxygen radicals spontaneously and this process is augmented by hypoxia/reoxygenation. Cu/Zn superoxide dismutase (SOD-1) is an important enzyme in cellular oxygen metabolism. To determine whether alterations in SOD-1 activity affect angiogenesis we used transgenic SOD-1 (Tg-SOD) mice with elevated level of SOD-1. Angiogenesis induced subcutaneously by bFGF in Tg-SOD mice was 3-fold higher than in control non-transgenic (ntg) mice. Oral administration of disulfiram (DSF), an inhibitor of SOD-1, inhibited angiogenesis in Tg-SOD mice as well as in CD1 nude mice. Effects of DSF on cultured cells were also tested. Application of DSF to cultured bovine capillary endothelial (BCE) cells caused inhibition of DNA synthesis and induction of apoptosis. These effects were prevented by addition of antioxidants, further indicating involvement of reactive oxygen species. DSF also reduced the level of glutathione and the production of H(2)O(2) in BCE cells. Moreover, PC12-SOD cells with elevated SOD-1 were less sensitive to DSF treatment then control cells. These data indicate that the effects of DSF are mediated by inhibition of SOD-1 activity. Tumor development is known to largely depend on angiogenesis. We found that oral administration of DSF to mice caused significant inhibition of C6 glioma tumor development and marked reduction (by 10-19-fold) in metastatic growth of Lewis lung carcinoma. The data suggest a role for SOD-1 in angiogenesis, establish DSF as a potential inhibitor of angiogenesis and raise the possibility that attenuating SOD-1 activity may be important in treatment of angiogenesis-dependent pathologies.